Mammalian layer 5 neurons initially exhibit exuberant subcortical projection, and many axonal branches are selectively pruned during development resulting in the area-specific projections seen in mature animals. This proposal outlines experiments that take advantage of the pruning defect observed in Otxl mutant mice to explore the cellular and molecular mechanisms underlying axon pruning. Mice with mutated Oixl alleles (by inserting EGFP and tau-LacZ reporter cassettes) will be generated. The first aim is to directly observe the axon pruning process in EGFP and tau-b-galactosidase-tagged neurons. The second aim is to determine whether Otxl functions cell-autonomously in axon pruning. The final aim is to study the molecular mechanism of axon pruning by identifying genes that are differentially regulated in Otxl mutant mice. These studies will further our understanding of how specific patterns of axonal connectivity are achieved through axon pruning.